title: Characterization of a Viral EPS-Depolymerase, a Potential Tool for Control of Fire Blight.
abstract: ABSTRACT A 3.3-kb fragment of genomic DNA from bacteriophage Phi-Ea1h encoding an amylovoran-directed depolymerase lyase was sequenced, and three open reading frames (ORFs) were detected. The first ORF could encode a lysozyme and the second a holin that may form a pore supporting cell lysis by the lysozyme. The third ORF encodes a protein of 657 amino acids and deletion mutation in this DNA fragment abolished extracellular polysaccharide (EPS)-degrading activity. A putative promoter and a ribosome binding sequence were located in front of the gene. A polymerase chain reaction product spanning the gene was inserted into multi copy plasmids including fusions with a Histidine-tagged sequence to facilitate its purification on a nickel nitrilotriacetic acid column. Maximal activity of the purified protein was observed between pH 4 and 5 at 52 degrees C. Visualized by staining with fluorescein isothiocyanate-labeled lectin from Abrus precatorious, the enzyme degraded the EPS-capsules of Erwina amylovora. In virulence assays, no symptoms were detected for a low inoculum of an E. amylovora strain when pear slices were soaked in a solution of depolymerase in contrast to control slices without addition of the enzyme. Furthermore, gfp- or lux-labeled E. amylovora cells were not propagated, when their amylovoran capsules were removed by the depolymerase. The enzyme could be a tool for biological control of fire blight.
authors.forename: W S
authors.initials: WS
authors.lastname: Kim
authors.forename: K
authors.initials: K
authors.lastname: Geider